Functional Characterization of the Gene Cluster from Pseudomonas syringae pv. phaseolicola NPS3121 Involved in Synthesis of Phaseolotoxin
Functional Characterization of the Gene Cluster from Pseudomonas syringae pv. phaseolicola NPS3121 Involved in Synthesis of Phaseolotoxin
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DOI：
10.1128/jb.01845-06
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发表时间：
2007-04-01
影响因子：
3.2
通讯作者：
Alvarez-Morales, Ariel
中科院分区：
文献类型：
--
作者：
Aguilera, Selene;Lopez-Lopez, Karina;Alvarez-Morales, Ariel
紫丁香假单胞菌菜豆白粉病是引起菜豆晕疫病的病原，它的特征是被水浸泡的病斑被一种被称为菜豆毒素的非寄主专性毒素作用而形成的绿晕包围。这种植物毒素抑制参与精氨酸生物合成的鸟氨酸氨基甲酰转移酶。不同的证据表明，参与相藻毒素产生的基因是聚集的。我们的实验室以前已经在这个簇中发现了两个基因：argK和AMTA，前者参与细菌对自身毒素的免疫，后者参与高精氨酸的合成。我们测定了紫丁香P.syringae PV中的argK和AMTA附近区域的序列。用来确定推测的菜豆毒素基因簇的限度，并确定组成它的基因的转录模式。我们报道的PHT簇由23个基因组成，其两侧是插入序列和转座酶。该簇内14个基因的突变导致其中11个基因的Tox(-)表型，而3个突变体表现出低水平的毒素产生。对选定的DNA片段与uidA、Northern Prospecing和逆转录-PCR的融合分析表明，存在五个转录单位，两个单顺反子和三个多顺反子；一个位于较大操纵子的内部。已经确定了每个启动子的转录起始位置，并确定了可能的启动子区域。初步结果还表明，phtL的基因产物参与了藻毒素的合成调控。
Pseudomonas syringae pv. phaseolicola is the causal agent of halo blight disease of beans (Phaseolus vulgaris L.), which is characterized by water-soaked lesions surrounded by a chlorotic halo resulting from the action of a non-host-specific toxin known as phaseolotoxin. This phytotoxin inhibits the enzyme ornithine carbamoyltransferase involved in arginine biosynthesis. Different evidence suggested that genes involved in phaseolotoxin production were clustered. Two genes had been previously identified in our laboratory within this cluster: argK, which is involved in the immunity of the bacterium to its own toxin, and amtA, which is involved in the synthesis of homoarginine. We sequenced the region around argK and amtA in P. syringae pv. phaseolicola NPS3121 to determine the limits of the putative phaseolotoxin gene cluster and to determine the transcriptional pattern of the genes comprising it. We report that the phaseolotoxin cluster (Pht cluster) is composed of 23 genes and is flanked by insertion sequences and transposases. The mutation of 14 of the genes within the cluster lead to a Tox(-) phenotype for 11 of them, while three mutants exhibited low levels of toxin production. The analysis of fusions of selected DNA fragments to uidA, Northern probing, and reverse transcription-PCR indicate the presence of five transcriptional units, two monocistronic and three polycistronic; one is internal to a larger operon. The site for transcription initiation has been determined for each promoter, and the putative promoter regions were identified. Preliminary results also indicate that the gene product of phtL is involved in the regulation of the synthesis of phaseolotoxin.